Plants are constantly challenged by environmental stress factors that affect their survival. Adverse biotic and abiotic stress factors can reduce the growth and productivity of plant crops. As tolerance to biotic and abiotic stresses has a direct impact on plant productivity (yield and product quality), mechanisms for conferring or enhancing stress tolerance have been widely studied and various approaches for conferring environmental stress tolerance have been described in the art. However, while most studied mechanisms are on either biotic or abiotic stress interactions, in nature plants respond and resist multiple stress factors.
Environmental stress factors have been estimated to cause depreciation in crop yield up to 70% when compared to the yield under favorable conditions (Boyer, Science 218, 443-448, 1982). Stability of crops to changes in environmental factors are therefore one of the most valued traits for breeding. However, traditional breeding is thwarted by the complexity of stress tolerance traits, low genetic variance of yield components and the lack of efficient selection techniques. It might therefore be useful to follow specific genes coding for stress tolerance components, in breeding by marker-assisted selection as well as by genetically engineering plants to be more stress tolerant.
Amidst the complexities of environmental stress reactions in crop plants the use of the simple model Arabidopsis, offers an opportunity for the precise genetic analysis of stress reaction pathways common to most plants. The relevance of the Arabidopsis model is evident in recent examples of improving drought, salt and freezing tolerance (Jaglo-Ottosen et al., Science 280, 104-106, 1998; Kasuga et al., Nat. Biotechnol. 17, 287-291, 1999) using genes identified in Arabidopsis. These genes are transcription factors of the ERF/AP2 family that regulate the expression of a number of downstream genes conferring stress resistance in a number of heterologous plants.
One of the most serious abiotic stresses plants have to cope with world-wide is drought stress or dehydration stress. Four-tenths of the world's agricultural land lies in arid or semi-arid regions. Apart from that, also plants grown in regions with relatively high precipitation may suffer spells of drought throughout the growing season. Many agricultural regions, especially in developing countries, have consistently low rain-fall and rely on irrigation to maintain yields. Water is scarce in many regions and its value will undoubtedly increase with global warming, resulting in an even greater need for drought tolerant crop plants, which maintain yield levels (or even have higher yields) and yield quality under low water availability. It has been estimated that the production of 1 kg of cotton requires about 15,000 liters of water in irrigated agriculture, while 1 kg of rice requires 4000 liters. Enhancing or engineering the tolerance of crop plants to short and long spells of drought and reducing the water requirement of crops grown in irrigated agriculture is clearly an important objective.
Although breeding (e.g. marker assisted) for drought tolerance is possible and is being pursued for a range of crop species (mainly cereals, such as maize, upland rice, wheat, sorghum, pearl millet, but also in other species such as cowpea, pigeon pea and Phaseolus bean), it is extremely difficult and tedious because drought tolerance or resistance is a complex trait, determined by the interaction of many loci and gene-environment interactions. Single, dominant genes, which confer or improve drought tolerance and which can be easily transferred into high yielding crop varieties and breeding lines are therefore sought after. Most water is lost through the leaves, by transpiration, and many transgenic approaches have focused on modifying the water loss through changing the leaves. For example WO00/73475 describes the expression of a C4 NADP+-malic enzyme from maize in tobacco epidermal cells and guard cells, which, according to the disclosure, increases water use efficiency of the plant by modulating stomatal aperture. Other approaches involve, for example, the expression of osmo-protectants, such as sugars (e.g. trehalose biosynthetic enzymes) in plants in order to increase water-stress tolerance, see e.g. WO99/46370. Yet other approaches have focused on changing the root architecture of plants.
To date another promising approach to enhance drought tolerance is the overexpression of CBF/DREB genes (DREB refers to dehydration response element binding; DRE binding), encoding various AP2/ERF (ethylene response factor) transcription factors (WO98/09521). Overexpression of the CBF/DREB1 proteins in Arabidopsis resulted in an increase in freezing tolerance (also referred to as freeze-induced dehydration tolerance) (Jaglo-Ottosen et al., Science 280, 104-106, 1998; Liu et al., Plant Cell 10, 1391-1406, 1998; Kasuga et al., Nat. Biotechnol. 17, 287-291, 1999; Gilmour et al. Plant Physiol. 124, 1854-1865, 2000) and enhanced the tolerance of the recombinant plants to dehydration caused either by water deficiency or exposure to high salinity (Liu et al., 1998, supra; Kasuga et al., 1999, supra). Another CBF transcription factor, CBF4, has been described to be a regulator of drought adaptation in Arabidopsis (Haake et al. 2002, Plant Physiology 130, 639-648).
Despite the availability of some genes which have been shown to enhance drought tolerance in a number of plant species, such as Brassicaceae and Solanaceae, there is a need for the identification of other genes with the ability to confer or improve drought tolerance when expressed in crop plants. In one embodiment, the present invention provides a new gene and associated mechanism which fulfil this need.
Biotic stresses like pathogen (bacteria, fungi, virus) or pests (insect, nematode) are the most common and a number of mechanisms normally protect plants against most of these threats. However, in certain cases plants display a susceptible reaction to specific pathogens or pests and are considered as host for the particular pathogen or pest. The host-pathogen interaction has been well characterized by the gene-for-gene concept where specific genes from the host plant and a pathogen/pest interact to either display a susceptible or a resistant reaction. Though the molecular genetics of such interactions have been well characterized in recent years, the use of such simple resistance genes has been hampered by the versatile mutability of the pathogen system that produces the diversity to overcome the resistance genes. In general the resistance genes belong to a few general classes of proteins that comprise of leucine rich repeats and additional domains. Though these genes and genetic interactions are interesting to study plant pathogen interactions, their employment for crop protection against a wider diversity and range of pathogens is still a ways away. Another way that can provide resistance is to use genes that are involved in the protection of plants to a diverse range of pathogens using mechanisms that do not rely on recognition of plant and pathogens. This would confer a non race specific resistance which is broader as it would confer resistance to a wider range of pathogens.